Objective:The aim is to establish regeneration system in vitro of Phytolacca acinosa Roxb. Methods: To screen regeneration system scheme of Phytolacca acinosa Roxb, young stem and stem node and leaves and petioles and apical buds of Phytolacca acinosa Roxb were chosen as explants, and MS was used as basic medium with different plant growth regulators combinations to induce callus and buds and roots. Results: Callus could be induced from apical buds and young stem with early emergence and high quality, and the callus induction rate reached 100% in medium MS+6-BA 0.5 mg/L+ 2.4-D 0.5 mg/L. Among them, only the callus produced from apical bud could differentiate cluster buds. The medium for bud differentiation was MS+6-BA 2.0 mg/L+NAA 0.25 mg/L, and the induction rate reached 98%. The suitable medium for rooting was 1/2 MS + NAA 0.3 mg/L with 100% induction rate. Conclusion:The regeneration system in vitro of Phytolacca acinosa Roxb was established and perfected, which laid a foundation for the construction of genetic transformation system in Phytolacca acinosa Roxb.